[ccp4bb] AW: [ccp4bb] Unknown positive electron density

2017-08-22 Thread Herman . Schreuder 
Dear Betty,

You have very high resolution, which helps you to identify your ligand, but the 
ligand may be disordered…
What I would do is to place some dummy atoms (e.g. waters) and refine and look 
if the molecule gets clearer. By scrolling the density in coot, you can 
identify the positions with the highest electron density were you should put 
your dummy atoms.

Looking at your pictures, as far as is possible without the ability to rotate 
them, I would try to fit deoxyribose-phosphate, or even a complete nucleotide. 
Maybe your prep contained some unreacted nucleotides and your anomalous peak is 
phosphorus.

Good luck!
Herman

Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Betty Chu
Gesendet: Montag, 21. August 2017 17:19
An: CCP4BB@JISCMAIL.AC.UK
Betreff: [ccp4bb] Unknown positive electron density

Dear ccp4bb,
I am refining a 1.40 Angstrom data set for a DNA oligonucleotide. While the 
model for the DNA fits very well into the density, there is a patch of positive 
electron density in the solvent space that we are having trouble with.

The screenshot can be viewed through this link:
https://cbsostorage.chem.umd.edu/owncloud/index.php/s/J5cKnOpCC4vb1VC

In the screenshot, the yellow color is the anomalous map and a barium ion is 
fitted into density near the positive green electron density.

The oligonucleotide was purchased from IDT. The crystallization condition is 
15% MPD, 120 mM BaCl2, and 30 mM NaCaC pH 6.4. I have tried modelling Ba2+ with 
coordinated waters, MPD, and sodium cacodylate into the electron density, but 
none of those fit well.
Any suggestions regarding the identity of this electron density is much 
appreciated. Thank you!

Sincerely,
Betty Chu
Paukstelis Research Group
Department of Chemistry and Biochemistry
University of Maryland, College Park

Re: [ccp4bb] AW: [ccp4bb] Unknown positive electron density

2017-08-22 Thread Pradeep Pallan 
Hi Betty,
Could you post a few more screen shots of different orientations of the
electron density?

If you see waters that are 1.6, 1.9 and 2.2 A away from the metal ion, I
would try
Mg2+ ion (trace amount of metal ions could be present in salt
solutions/buffers as an impurity)
and refine it (although 1.6Å is too close). In general, Mg2+ ions would
show an octahedral
geometry.





On Tue, Aug 22, 2017 at 3:35 AM,  wrote:

> Dear Betty,
>
>
>
> You have very high resolution, which helps you to identify your ligand,
> but the ligand may be disordered…
>
> What I would do is to place some dummy atoms (e.g. waters) and refine and
> look if the molecule gets clearer. By scrolling the density in coot, you
> can identify the positions with the highest electron density were you
> should put your dummy atoms.
>
>
>
> Looking at your pictures, as far as is possible without the ability to
> rotate them, I would try to fit deoxyribose-phosphate, or even a complete
> nucleotide. Maybe your prep contained some unreacted nucleotides and your
> anomalous peak is phosphorus.
>
>
>
> Good luck!
>
> Herman
>
>
>
> *Von:* CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *Im Auftrag von
> *Betty Chu
> *Gesendet:* Montag, 21. August 2017 17:19
> *An:* CCP4BB@JISCMAIL.AC.UK
> *Betreff:* [ccp4bb] Unknown positive electron density
>
>
>
> Dear ccp4bb,
>
> I am refining a 1.40 Angstrom data set for a DNA oligonucleotide. While
> the model for the DNA fits very well into the density, there is a patch of
> positive electron density in the solvent space that we are having trouble
> with.
>
> The screenshot can be viewed through this link:
> https://cbsostorage.chem.umd.edu/owncloud/index.php/s/J5cKnOpCC4vb1VC
> 
>
> In the screenshot, the yellow color is the anomalous map and a barium ion
> is fitted into density near the positive green electron density.
>
>
> The oligonucleotide was purchased from IDT. The crystallization condition
> is 15% MPD, 120 mM BaCl2, and 30 mM NaCaC pH 6.4. I have tried modelling
> Ba2+ with coordinated waters, MPD, and sodium cacodylate into the electron
> density, but none of those fit well.
>
> Any suggestions regarding the identity of this electron density is much
> appreciated. Thank you!
>
>
>
> Sincerely,
>
> Betty Chu
>
> Paukstelis Research Group
>
> Department of Chemistry and Biochemistry
>
> University of Maryland, College Park
>



-- 

---
Pradeep Pallan

Re: [ccp4bb] AW: [ccp4bb] Unknown positive electron density

2017-08-24 Thread Betty Chu 
Thank you all for your replies and suggestions.

I have fitted in some dummy waters as Herman suggested and this is the
result after one round of refinement. The waters are positioned where there
is highest electron density, but I am not sure how to move forward from
here.

https://cbsostorage.chem.umd.edu/owncloud/index.php/s/VTQ6Uinp1UVIWGR
https://cbsostorage.chem.umd.edu/owncloud/index.php/s/NUZKClayzSMn7PP
https://cbsostorage.chem.umd.edu/owncloud/index.php/s/0FZhxtgcQhhqnaA

I have also tried fitting in a nucleotide, but the geometry of it does not
fit the density well. We do not think this is due to Fourier truncation
ripples since the positive density is not at the crystallographic symmetry
axis.

Betty

On Tue, Aug 22, 2017 at 10:34 AM, Pradeep Pallan 
wrote:

> Hi Betty,
> Could you post a few more screen shots of different orientations of the
> electron density?
>
> If you see waters that are 1.6, 1.9 and 2.2 A away from the metal ion, I
> would try
> Mg2+ ion (trace amount of metal ions could be present in salt
> solutions/buffers as an impurity)
> and refine it (although 1.6Å is too close). In general, Mg2+ ions would
> show an octahedral
> geometry.
>
>
>
>
>
> On Tue, Aug 22, 2017 at 3:35 AM,  wrote:
>
>> Dear Betty,
>>
>>
>>
>> You have very high resolution, which helps you to identify your ligand,
>> but the ligand may be disordered…
>>
>> What I would do is to place some dummy atoms (e.g. waters) and refine and
>> look if the molecule gets clearer. By scrolling the density in coot, you
>> can identify the positions with the highest electron density were you
>> should put your dummy atoms.
>>
>>
>>
>> Looking at your pictures, as far as is possible without the ability to
>> rotate them, I would try to fit deoxyribose-phosphate, or even a complete
>> nucleotide. Maybe your prep contained some unreacted nucleotides and your
>> anomalous peak is phosphorus.
>>
>>
>>
>> Good luck!
>>
>> Herman
>>
>>
>>
>> *Von:* CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *Im Auftrag
>> von *Betty Chu
>> *Gesendet:* Montag, 21. August 2017 17:19
>> *An:* CCP4BB@JISCMAIL.AC.UK
>> *Betreff:* [ccp4bb] Unknown positive electron density
>>
>>
>>
>> Dear ccp4bb,
>>
>> I am refining a 1.40 Angstrom data set for a DNA oligonucleotide. While
>> the model for the DNA fits very well into the density, there is a patch of
>> positive electron density in the solvent space that we are having trouble
>> with.
>>
>> The screenshot can be viewed through this link:
>> https://cbsostorage.chem.umd.edu/owncloud/index.php/s/J5cKnOpCC4vb1VC
>> 
>>
>> In the screenshot, the yellow color is the anomalous map and a barium ion
>> is fitted into density near the positive green electron density.
>>
>>
>> The oligonucleotide was purchased from IDT. The crystallization condition
>> is 15% MPD, 120 mM BaCl2, and 30 mM NaCaC pH 6.4. I have tried modelling
>> Ba2+ with coordinated waters, MPD, and sodium cacodylate into the electron
>> density, but none of those fit well.
>>
>> Any suggestions regarding the identity of this electron density is much
>> appreciated. Thank you!
>>
>>
>>
>> Sincerely,
>>
>> Betty Chu
>>
>> Paukstelis Research Group
>>
>> Department of Chemistry and Biochemistry
>>
>> University of Maryland, College Park
>>
>
>
>
> --
>
> ---
> Pradeep Pallan
>